CST Guide:

Section I: Research Areas
SAPK/JNK Signaling
Growth Factors, UV
RTKs
CrkII
CrkL
Shc
GRB2
HPK1
Cytoplasm
Nucleus
PI3K
14-3-3
SOS
MAPKKKs :
TAK1
Ras
MKK4/7
JNK1/2/3
IRS-1
c-Abl 14-3-3 14-3-3
DUSPs
c-Abl
Cellular Stress,
γ Radiation
Rac
MEKK1/4
MLK2/3
DLK
TpI-2
TAO1/2
NO
Cdc42
Rho
MLKs
POSH
JNK
JNK
FasL, UV
Inflammatory Cytokines
GCK
ASK1
Rac
c-Jun ATF-2 Elk-1 Smad4 p53 NFAT4 NFATc1 Stat3
TRADD
TRAF2
JIP1,2,3
RIP
Daxx
Transcription
Oxidative
Stress
Apoptosis
Cdc42
MAPKKKs
Bax
HSF1
Gα 12/13
14-3-3
Bax
Growth
Differentiation
Survival
Apoptosis
G 12/13 -coupled
Receptors
Gβγ
PI3Kγ
Mitochondrial
Translocation/
Apoptosis
Stress-activated protein kinases (SAPK)/Jun amino-terminal kinases (JNK) are members of the MAPK family and are activated by a variety of environmental stresses, inflammatory
cytokines, growth factors, and GPCR agonists. Stress signals are delivered to this cascade by small GTPases of the Rho family (Rac, Rho, cdc42). As with the other
MAPKs, the membrane proximal kinase is a MAPKKK, typically MEKK1–4, or a member of the mixed lineage kinases (MLK) that phosphorylates and activates MKK4 (SEK) or
MKK7, the SAPK/JNK kinases. Alternatively, MKK4/7 can be activated by a member of the germinal center kinase (GCK) family in a GTPase-independent manner. SAPK/JNK
translocates to the nucleus where it can regulate the activity of multiple transcription factors.
Select Reviews:
Bogoyevitch, M.A., Ngoei, K.R., Zhao, T.T., Yeap, Y.Y., and Ng, D.C. (2010) Biochim. Biophys. Acta. 1804, 463–475. • Chen, F. (2012) Cancer Res. 72, 379–386. • Davies,
C. and Tournier, C. (2012) Biochem. Soc. Trans. 40, 85–89. • Engström, W., Ward, A., and Moorwood, K. (2010) Cell Prolif. 43, 56–66. • Haeusgen, W., Herdegen, T., and
Waetzig, V. (2011) Eur. J. Cell Biol. 90, 536–544. • Verma, G., and Datta, M. (2012) J. Cell Physiol. 227, 1791–1795.
PI3 Kinase/Akt Signaling
Activation of Akt
The serine/threonine kinase Akt/PKB exists as three isoforms in mammals. Akt1 and Akt2 have a wide
tissue distribution, whereas Akt3 is expressed in testes and brain. Akt regulates multiple biological processes
including cell survival, proliferation, growth, and glycogen metabolism. Various growth factors,
hormones, and cytokines activate Akt by binding their cognate receptor tyrosine kinase (RTK), cytokine
receptor, or G protein-coupled receptor (GPCR) and triggering activation of the lipid kinase PI3K, which
generates PIP 3 at the plasma membrane. Akt binds PIP 3 through its pleckstrin homology (PH) domain,
resulting in translocation of Akt to the membrane. Akt is activated through a dual phosphorylation
mechanism. PDK1, which is also brought to the membrane through its PH domain, phosphorylates Akt
within its activation loop at Thr308. A second phosphorylation at Ser473 within the C-terminus is also
required for activity and is carried out by the mTOR-Rictor complex, mTORC2.
Insulin stimulation results in clustering of phospho-Akt at the membrane.
Phospho-Akt (Thr308) (D25E6) XP ® Rabbit
mAb #13038: Confocal IF analysis of C2C12
cells, insulin-treated (100 nM, 15 min; left) or
treated with LY294002 #9901 (50 μM, 2 hr;
right), using #13038 (green). Actin filaments
were labeled with DY-554 phalloidin (red). Blue
pseudocolor = DRAQ5 ® #4084 (fluorescent
DNA dye).
Growth factor stimulation results in phosphorylation
of Akt at Thr308, one of two phosphorylations
necessary for Akt activation.
Phospho-Akt (Thr308) (D25E6) XP ® Rabbit mAb #13038: Western blot analysis of
extracts from NIH/3T3 cells, untreated or treated with Human Platelet-Derived Growth
Factor AA (hPDGF-AA) #8913 (100 ng/ml, 5 min; +), and untreated LNCaP and PC-3
cells, using #1308 (upper) or Akt (pan) (C67E7) Rabbit mAb #4691 (lower).
Lanes
1. NIH/3T3
2. NIH/3T3
3. LNCaP
4. PC-3
kDa
140
100
80
60
50
60
50
1 2 3 4
Phospho-
Akt (Thr308)
Akt (pan)
– + – – hPDGF
Inhibition of Akt Activity
PTEN, a lipid phosphatase that catalyzes the dephosphorylation of PIP 3 , is a major negative regulator of
Akt signaling. Loss of PTEN function has been implicated in many human cancers. Akt activity is also
negatively regulated by the phosphatases PP2A and PHLPP, as well as by the chemical modulators
wortmannin and LY294002, both of which are inhibitors of PI3K.
PTEN expression in human colon
PTEN (D4.3) XP ® Rabbit mAb #9188: IHC analysis
of paraffin-embedded human colon using #9188.
kDa
100
chapter 02: Signaling
Akt-activating treatments
and absence
of PTEN result in
phosphorylation of
Akt at Ser473.
A
80
60
50
80
60
50
PC-3
– +
– +
– –
NIH/3T3
– –
– –
– +
Phospho-Akt (Ser473)
is expressed in human
lung carcinoma.
B
Phospho-
Akt (Ser473)
Akt
wortmannin
LY294002
PDGF
Phospho-Akt (Ser473) (D9E) XP ®
Rabbit mAb #4060: WB analysis of
extracts from PC-3 cells (PTEN null),
untreated or treated with LY294002
#9901/Wortmannin #9951, and NIH/3T3
cells, serum-starved or PDGF-treated
(A), using #4060 (upper) or Akt (pan)
(C67E7) Rabbit mAb #4691 (lower). IHC
analysis of paraffin-embedded human
lung carcinoma (B) using #4060.
© 2002–2015 Cell Signaling Technology, Inc. • We would like to thank Prof. John Blenis, Harvard Medical School, Boston, MA, for reviewing this diagram.
48 For Research Use Only. Not For Use in Diagnostic Procedures. See pages 302 & 303 for Pathway Diagrams, Application, and Reactivity keys.
www.cellsignal.com/cstakt 49